Use of synthetic peptides to delineate discontinuous sequence regions involved in epitope sites of the thyrotropin β-subunit

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Abstract

In this investigation, an overlapping set of synthetic peptides spanning the entire primary structures of the β-subunit of bovine and human thyrotropin, bTSHβ and hTSHβ respectively, have been prepared to aid the delineation of the amino acid sequence regions involved in two spatially related epitopes of bTSH. These peptides were then evaluated for their ability to inhibit the binding of two anti-hTSH monoclonal antibodies, designated mAb279 and mAb299, to radiolabeled I125-bTSHβ using competitive radioimmunoassay procedures. Synthetic peptides related to the sequence region b/hTSHβ [56-68] were found to specifically inhibit the binding of I 125-bTSHβ to mAb299, whilst having no effect on the binding of mAb279. In previous studies we have shown that mAb279 and mAb299 recognise epitopic sites located within the receptor-binding site of the TSH β-subunit. This investigation has therefore permitted identification of a contribution to the receptor binding site from the TSHβ [56-68] sequence, which forms part of the L3 loop region of the TSH β-subunit that is held in close proximity to the L1 loop region and the C-terminus of the TSH β-subunit by the disulphide bonds TSHβ[Cys16-Cys 67] and TSHβ[Cys19-Cys105]. This finding is in agreement with previous investigations which have shown that TSHβ [Tyr59] and TSHβ [Tyr74] are also associated with the mAb299 epitope site, as well as contributing to the receptor binding region of the TSH β-subunit.

Original languageEnglish
Pages (from-to)185-192
Number of pages8
JournalInternational Journal of Peptide Research and Therapeutics
Volume6
Issue number2-3
DOIs
Publication statusPublished - 1 Jan 1999

Keywords

  • Discontinuous epitopes
  • Glycoprotein hormones
  • Monoclonal antibodies
  • Receptor binding site
  • Synthetic peptides

Cite this

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title = "Use of synthetic peptides to delineate discontinuous sequence regions involved in epitope sites of the thyrotropin β-subunit",
abstract = "In this investigation, an overlapping set of synthetic peptides spanning the entire primary structures of the β-subunit of bovine and human thyrotropin, bTSHβ and hTSHβ respectively, have been prepared to aid the delineation of the amino acid sequence regions involved in two spatially related epitopes of bTSH. These peptides were then evaluated for their ability to inhibit the binding of two anti-hTSH monoclonal antibodies, designated mAb279 and mAb299, to radiolabeled I125-bTSHβ using competitive radioimmunoassay procedures. Synthetic peptides related to the sequence region b/hTSHβ [56-68] were found to specifically inhibit the binding of I 125-bTSHβ to mAb299, whilst having no effect on the binding of mAb279. In previous studies we have shown that mAb279 and mAb299 recognise epitopic sites located within the receptor-binding site of the TSH β-subunit. This investigation has therefore permitted identification of a contribution to the receptor binding site from the TSHβ [56-68] sequence, which forms part of the L3 loop region of the TSH β-subunit that is held in close proximity to the L1 loop region and the C-terminus of the TSH β-subunit by the disulphide bonds TSHβ[Cys16-Cys 67] and TSHβ[Cys19-Cys105]. This finding is in agreement with previous investigations which have shown that TSHβ [Tyr59] and TSHβ [Tyr74] are also associated with the mAb299 epitope site, as well as contributing to the receptor binding region of the TSH β-subunit.",
keywords = "Discontinuous epitopes, Glycoprotein hormones, Monoclonal antibodies, Receptor binding site, Synthetic peptides",
author = "Gomme, {Peter T.} and Thompson, {Philip E.} and James Whisstock and Stanton, {Peter G.} and Hearn, {Milton T.W.}",
year = "1999",
month = "1",
day = "1",
doi = "10.1007/BF02443635",
language = "English",
volume = "6",
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journal = "International Journal of Peptide Research and Therapeutics",
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TY - JOUR

T1 - Use of synthetic peptides to delineate discontinuous sequence regions involved in epitope sites of the thyrotropin β-subunit

AU - Gomme, Peter T.

AU - Thompson, Philip E.

AU - Whisstock, James

AU - Stanton, Peter G.

AU - Hearn, Milton T.W.

PY - 1999/1/1

Y1 - 1999/1/1

N2 - In this investigation, an overlapping set of synthetic peptides spanning the entire primary structures of the β-subunit of bovine and human thyrotropin, bTSHβ and hTSHβ respectively, have been prepared to aid the delineation of the amino acid sequence regions involved in two spatially related epitopes of bTSH. These peptides were then evaluated for their ability to inhibit the binding of two anti-hTSH monoclonal antibodies, designated mAb279 and mAb299, to radiolabeled I125-bTSHβ using competitive radioimmunoassay procedures. Synthetic peptides related to the sequence region b/hTSHβ [56-68] were found to specifically inhibit the binding of I 125-bTSHβ to mAb299, whilst having no effect on the binding of mAb279. In previous studies we have shown that mAb279 and mAb299 recognise epitopic sites located within the receptor-binding site of the TSH β-subunit. This investigation has therefore permitted identification of a contribution to the receptor binding site from the TSHβ [56-68] sequence, which forms part of the L3 loop region of the TSH β-subunit that is held in close proximity to the L1 loop region and the C-terminus of the TSH β-subunit by the disulphide bonds TSHβ[Cys16-Cys 67] and TSHβ[Cys19-Cys105]. This finding is in agreement with previous investigations which have shown that TSHβ [Tyr59] and TSHβ [Tyr74] are also associated with the mAb299 epitope site, as well as contributing to the receptor binding region of the TSH β-subunit.

AB - In this investigation, an overlapping set of synthetic peptides spanning the entire primary structures of the β-subunit of bovine and human thyrotropin, bTSHβ and hTSHβ respectively, have been prepared to aid the delineation of the amino acid sequence regions involved in two spatially related epitopes of bTSH. These peptides were then evaluated for their ability to inhibit the binding of two anti-hTSH monoclonal antibodies, designated mAb279 and mAb299, to radiolabeled I125-bTSHβ using competitive radioimmunoassay procedures. Synthetic peptides related to the sequence region b/hTSHβ [56-68] were found to specifically inhibit the binding of I 125-bTSHβ to mAb299, whilst having no effect on the binding of mAb279. In previous studies we have shown that mAb279 and mAb299 recognise epitopic sites located within the receptor-binding site of the TSH β-subunit. This investigation has therefore permitted identification of a contribution to the receptor binding site from the TSHβ [56-68] sequence, which forms part of the L3 loop region of the TSH β-subunit that is held in close proximity to the L1 loop region and the C-terminus of the TSH β-subunit by the disulphide bonds TSHβ[Cys16-Cys 67] and TSHβ[Cys19-Cys105]. This finding is in agreement with previous investigations which have shown that TSHβ [Tyr59] and TSHβ [Tyr74] are also associated with the mAb299 epitope site, as well as contributing to the receptor binding region of the TSH β-subunit.

KW - Discontinuous epitopes

KW - Glycoprotein hormones

KW - Monoclonal antibodies

KW - Receptor binding site

KW - Synthetic peptides

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U2 - 10.1007/BF02443635

DO - 10.1007/BF02443635

M3 - Article

VL - 6

SP - 185

EP - 192

JO - International Journal of Peptide Research and Therapeutics

JF - International Journal of Peptide Research and Therapeutics

SN - 1573-3149

IS - 2-3

ER -